In mid-60s Rosenberg serendipitously discovered the anticancer properties of cisplatin (cis-dichlorodiamminoplatinum(II), cis-[PtIICl2(NH3)2]). Following the encouraging results of these studies, it was approved by the Food and Drug Administration (FDA) at the end of 1978 for the treatment of genitourinary tumors. At present, cisplatin is one of the most effective drugs used for the treatment of testicular and, in combination with other chemotherapeutic agents, of ovarian, small-cell lung, bladder, cervical, brain and breast cancer. However, all chemotherapeutic drugs have drawbacks, and cisplatin is no exception. In fact, in spite of its therapeutic success in the treatment of several types of tumors, its high effectiveness is severely hindered by some adverse side-effects such as nausea, alopecia, ototoxicity, neurotoxicity, myelosuppression, and nephrotoxicity. A second major drawback is tumor resistance, either acquired during cycles of therapy with cisplatin (occurring in patients with, for example, ovarian cancer) or intrinsic (observed in patients with, for example, colorectal, prostate, lung or breast cancer) [L. Kelland, Nat. Rev. Cancer 2007, 7, 573-584]. Thus, much of the early effort in the design of new platinum drugs was aimed at making cisplatin-based therapy safer to patients, in particular, lessening or removing unpredictable and severe side-effects, providing oral bioavailability, and overcoming both intrinsic and acquired resistance. Efforts to mitigate the drawbacks have prompted chemists to synthesize a variety of analogues, but only a handful of new drugs were shown to be suitable for clinical application, amongst which carboplatin (cis-(1,1-cyclobutanedicarboxylato)diamminoplatinum(II)), oxaliplatin (eyhanedithioato-O,O′)(R,R)-1,2-diaminocyclohexano-N,N)platinum(II)), and satraplatin (cis,trans, cis-dichlorobis(acetate-O)amino(cyclhexylamino)platinum (IV)).
Carboplatin is essentially devoid of nephrotoxicity, less neurotoxic, and less toxic to the gastrointestinal tract. By contrast, myelosuppression, principally thrombocytopenia, is its dose-limiting factor. Oxaliplatin is especially interesting for tumors which do not or hardly respond to cisplatin, for instance colorectal tumors. Nevertheless, neurotoxicity was proved to be its major drawback. Satraplatin, a Pt(IV) compound which is reduced in vivo to some Pt(II) analogues, was proved promising in terms of treatment regime since it can be administered without hospitalization.
As previously stated, kidney toxicity limits the use of cisplatin and related platinum-based therapeutics. Nephrotoxicity may result from either too high administered doses or accumulation of cisplatin in the body. The effects of cisplatin on renal functions are not completely understood, but recent research has provided new insights on the mechanism of cisplatin nephrotoxicity, especially on the signaling pathways leading to tubular cell death and inflammation. It has been hypothesized that renal failure may be induced by platinum binding to and inactivation of thiol-containing enzymes [N. Pabla, Z. Dong, Kidney Int 2008, 73, 994-1007]. Thus, a number of thiol-based and sulfur-containing nucleophiles have been tested as chemoprotectants to modulate cisplatin nephrotoxicity [R. T. Dorr, “A review of the modulation of cisplatin toxicities by chemoprotectants” in: H. M. Pinedo, J. H. Schornagel (Eds.), Platinum and Other Metal Coordination Compounds in Cancer Chemotherapy 2, Plenum Press, New York, 1996, pp. 131-154]. Two main issues have to take into account for the development of chemoprotectants: (i) the selective protection of non-tumor normal tissues, and (ii) the addition of little, if any, toxicity. Many sulfur-based chemoprotectants such as L-BSO (L-buthionine sulfoximine), disulfiram (or antabuse, tetraethylthiuram disulfide), NAC (N-acetylcysteine), mesna (S-mercaptoethane sulfonate sodium salt), sodium thiosulfate, and ORG-2766 (a melanocortin-derived peptide) have been tested to modulate cisplatin renal toxicity, and several showed promising for clinical use. However, a selective protection of normal tissues without inhibition of antitumor effects was proved challenging.
In this regard, positive outcomes were obtained with sodium diethyldithiocarbamate (DEDTNa, Na((CH3CH2)2NCSS)). In fact, it was shown to provide protection against renal, gastrointestinal and bone marrow toxicity induced by cisplatin without decreasing its antitumor property. Its chemoprotective effect results from the capability to remove platinum from thiol groups of proteins without reversal of platinum-DNA adducts, responsible for its antitumor activity. Platinum-DNA adducts were shown to decrease by ca. 50% when cells were treated with DEDTNa soon after cisplatin administration, thus causing a loss of therapeutic effect, whereas no change in anticancer activity was observed when DEDTNa was administered 3 h after cisplatin.
However, the overall nephroprotective benefits of DEDTNa are significantly limited by the acute toxicity profile of dithiocarbamates themselves. In fact, potential human health hazards associated with free (i.e. not coordinated) dithiocarbamates is still being investigated, including genotoxicity and possible carcinogenicity [R. T. Dorr et al. 1996, ref. cit.].
Nevertheless, the wide success of platinum drugs promoted the development of both alternative platinum and other metal-based compounds that, at least in principle, might resemble its anticancer behavior.
Based on these assumptions, Faraglia G. and co-workers [G. Faraglia, D. Fregona, S. Sitran, L. Giovagnini, C. Marzano, F. Baccichetti, U. Casellato, R. Graziani, J. Inorg. Biochem. 2001, 83, 31-40] have previously designed mixed dithiocarbamato/amino Pt(II) and Pd(II) complexes ([MII(μ-Cl)(ESDT)]2, M=Pt and Pd, ESDT=CH3CH2O(O)CCH2N(CH3)CSS; [MIICl(ESDT)(py)], M=Pt and Pd, ESDT=CH3CH2O(O)CCH2N(CH3)CSS, py=pyridine; [MIICl(ESDT)(n-pra)], M=Pd, ESDT=CH3CH2O(O)CCH2N(CH3)CSS, n-pra=n-propylamine; [MIICl(ESDT)(en)]Cl, M=Pt and Pd, ESDT=CH3CH2O(O)CCH2N(CH3)CSS, en=ethylenediamine; [MIICl(ESDT)(n-pra)2]Cl, M=Pd, ESDT=CH3CH2O(O)CCH2N(CH3)CSS, n-pra=n-propylamine) potentially able to combine the cytotoxic activity of the metal centers with lack of nephrotoxicity. These species contain: (i) an amino ligand as most bioactive Pt(II) complexes, (ii) a good leaving group (chloride) which may undergo hydrolysis and bind DNA, thus resembling cisplatin mechanism of action, and (iii) an S,S′-chelating ligand (dithiocarbamate) potentially able to prevent interactions of the metal centre with sulfur-containing proteins, therefore reducing renal toxicity.
These complexes were evaluated for their in vitro cytotoxic activity toward human squamous cervical adenocarcinoma (HeLa) and human leukemic promyelocites (HL60) cells. Since complex [PtIICl(ESDT)(py)] showed the most promising cytotoxic properties, it was also evaluated on cisplatin-sensitive (2008) and—resistant (C13*) human ovarian carcinoma cells [C. Marzano, D. Fregona, F. Baccichetti, A. Trevisan, L. Giovagnini, F. Bordin, Chem. Biol. Interact. 2002, 140, 215-229]. Intriguingly, it induced a greater inhibition of tumor cell growth than cisplatin, and a complete lack of cross-resistance in C13* cells was observed. In addition, it was proved to be less efficient to platinate DNA, suggesting that its cytotoxic activity and the ability to overcome cisplatin resistance might be related to a different mechanism of interaction with DNA and/or with other key cellular components [C. Marzano et al. 2002, ref. cit.]. Finally, experimental results confirmed that [PtIICl(ESDT)(py)] was able to induce five-fold lower renal toxicity than cisplatin [C. Marzano, A. Trevisan, L. Giovagnini, D. Fregona, Toxicol. in Vitro 2002, 16, 413-419].
Despite these encouraging results, further evaluation of [PtIICl(ESDT)(py)] as potential anticancer agent was dismissed due to its poor water solubility and relatively low stability under physiological conditions. Fregona D. and co-workers carried out further studies on a number of analogous mixed dithiocarbamato/amino Pt(II) and Pd(II) complexes that exhibited high in vitro cytotoxicity together with low or even lack of nephrotoxicity [V. Alverdi, L. Giovagnini, C. Marzano, R. Seraglia, F. Bettio, S. Sitran, R. Graziani, D. Fregona, J. Inorg. Biochem. 2004, 98, 1117-1128], but long synthetic routes and inadequate stability in physiological environment was the major drawback.
In order to overcome these issues, Fregona D. and co-workers have recently explored alternative solutions, the most promising being the design of dithiocarbamato derivatives of metals other than platinum and palladium, such as Cu(II), Zn(II), and Ru(III) [L. Giovagnini, S. Sitran, M. Montopoli, L. Caparrotta, M. Corsini, C. Rosani, P. Zanello, Q. P. Dou, D. Fregona, Inorg. Chem. 2008, 47, 6336-6343; L. Giovagnini, S. Sitran, I. Castagliuolo, P. Brun, M. Corsini, P. Zanello, A. Zoleo, A. Maniero, B. Biondi, D. Fregona, Dalton Trans. 2008, 6699-6708].
In this context, Au(III) compounds are emerging as a new class of metal complexes with outstanding cytotoxic properties and are currently being evaluated as potential antitumor agents. Given their traditional use in medicine in the treatment of rheumatoid arthritis, gold compounds are a possible alternative to platinum drugs. In fact, their antiarthritic activity arises from the known immunosuppressive and antiinflammatory actions, thus establishing a connection, at least in principle, between the two therapies. Au(III) complexes show chemical features that are very close to those of clinically employed Pt(II) complexes, such as the preference for square-planar coordination and the typical d8 electronic configuration, making them very attractive for testing as antineoplastic drugs. Surprisingly, despite this strict similarity, little literature data exist on the use of Au(III) complexes as anticancer agents, the paucity of data probably deriving from their high redox potential and relatively poor stability, which make their use rather problematic under physiological conditions. Recently, Fregona D. et al. have reported on some Au(III)-dithiocarbamato derivatives of the type [AuIIIX2(dtc)] (X=Cl, Br; dtc=various dithiocarbamato ligands: MSDT=CH3O(O)CCH2N(CH3)CSS; ESDT=CH3CH2O(O)CCH2N(CH3)CSS; DMDT=(CH3)2NCSS), which have been designed in such a way to reproduce very closely the main features of cisplatin. From comparative in vitro cytotoxicity studies of Pt(II)-, Pd(II)-, and Au(III)-MSDT (MSDT=CH3O(O)CCH2N(CH3)CSS) derivatives on human squamous cervical adenocarcinoma (HeLa) and human leukemic promyelocites (HL60) cells, Au(III) complexes resulted to be significantly more active than both cisplatin and the Pt(II) and Pd(II) counter-parts under the same experimental conditions [L. Giovagnini, L. Ronconi, D. Aldinucci, D. Lorenzon, S. Sitran, D. Fregona, J. Med. Chem. 2005, 48, 1588-1595]. [AuIIIX2(MSDT)]-type compounds were proved to suppress, in a dose-dependent way, cell growth on a panel of acute myelogenous leukemia cell lines with IC50 values ca. 10-fold lower than the reference drug by inducing DNA fragmentation and cell apoptosis [D. Aldinucci, D. Lorenzon, L. Stefani, L. Giovagnini, A. Colombatti, D. Fregona, Anti-Cancer Drugs 2007, 18, 323-332]. On account of these encouraging results, Fregona D. and co-workers carried on the same way by developing other Au(III)-dithiocarbamato derivatives of the type [AuIIIX2(DMDT)] and [AuIIIX2(ESDT)] (X=Cl, Br; DMDT=(CH3)2NCSS; ESDT=CH3CH2O(O)CCH2N(CH3)CSS). These compounds were proved to be much more cytotoxic in vitro than cisplatin even towards human tumor cell lines intrinsically resistant to cisplatin itself. Moreover, they appeared to be much more active also on cisplatin-resistant cell lines, with activity levels comparable to those on the corresponding cisplatin-sensitive cell lines, ruling out the occurrence of cross-resistance phenomena [L. Ronconi, L. Giovagnini, C. Marzano, F. Bettio, R. Graziani, G. Pilloni, D. Fregona, Inorg. Chem. 2005, 44, 1867-1881].
Their behavior under physiological conditions and DNA binding properties have been also evaluated [L. Ronconi, C. Marzano, P. Zanello, M. Corsini, G. Miolo, C. Macca, A. Trevisan, D. Fregona, J. Med. Chem. 2006, 49, 1648-1657]. These Au(III) complexes showed high reactivity toward some biologically-relevant isolated macromolecules, resulting in a dramatic inhibition of both DNA and RNA synthesis and inducing DNA lesions with faster kinetics than cisplatin, supporting the hypothesis of a different mechanism of action compared to platinum drugs.
In this regard, the same research group have recently identified the proteasome as a major in vitro and in vivo target for these Au(III)-dithiocarbamato derivatives [V. Milacic, D. Chen, L. Ronconi, K. R. Landis-Piwowar, D. Fregona, Q. P. Dou, Cancer Res. 2006, 66, 10478-10486]. In particular, the authors showed that the inhibition of the proteasomal activity (especially, chymotrypsin-like activity) by [AuIIIBr2(DMDT)] is a strong apoptotic stimulus in the highly metastatic MDA-MB-231 breast cancer cell cultures and tumors. Fregona D. et al. also showed that treatment of MDA-MB-231 tumor-bearing nude mice with compound [AuIIIBr2(DMDT)] resulted in significant inhibition of tumor growth, as a consequence of proteasomal inhibition and apoptosis induction, together with lack of systemic toxicity, weight loss, decreased activity, or anorexia.
Saggioro D. et al. have extended the biological evaluation to mitochondria as potential target of complexes [AuIIICl2(DMDT)], [AuIIIBr2(DMDT)], [AuIIICl2(ESDT)] and [AuIIIBr2(ESDT)] and found that they induce cancer cell death through both apoptotic and non-apoptotic mechanisms [D. Saggioro, M. P. Rigobello, L. Paloschi, A. Folda, S. A. Moggach, S. Parsons, L. Ronconi, D. Fregona, A. Bindoli, Chem. Biol. 2007, 14, 1128-1139]. They also inhibit thioredoxin reductase activity, generate free radicals, modify some mitochondrial functions, and increase ERK1/2 phosphorylation. In vivo antitumor activity as well as tolerability and nephrotoxicity of [AuIIIBr2(ESDT)] have been evaluated and promising results were obtained. Thus, considering the potential advantages in terms of noticeable in vitro and in vivo antitumor activity, lack of cross-resistance with cisplatin and reduced adverse side-effects, this kind of Au(III)-dithiocarbamato derivatives may be regarded as prospective metal-based anticancer drugs [V. Milacic, D. Fregona, Q. P. Dou, Histol. Histopathol. 2008, 23, 101-108].
Notwithstanding these positive achieved outcomes, improvement of the therapeutic efficiency of this class of complexes is still an issue as far as the crossing of cell membrane is concerned.
Actually, cellular uptake of therapeutic agents is still a challenging task because of the plasma membrane, which constitutes an impermeable barrier for most of these molecules. In order to circumvent this issue, several carrier-mediated delivery systems have been developed. Among them, much attention has recently given to the use of peptide-based delivery systems. Peptide transporters are integral plasma membrane proteins that mediate the cellular uptake of di- and tri-peptides, and peptide-like drugs [A. Biegel, S. Gebauera, B. Hartrodta, I. Knütter, K. Neuberta, M. Brandschb, I. Thondorf, Eur. J. Pharm. Sci. 2007, 32, 69-76]. Two peptide transporters, namely PEPT1 and PEPT2, have been identified in mammals [Rubio-Aliaga, H. Daniel Trends Pharmacol. Sci. 2002, 23, 434-440]. They are present predominantly in epithelial cells of the small intestine, mammary glands, lung, choroid plexus and kidney, but are also found in other cell types. A unique feature is their capability for sequence-independent transport of all possible di- and tri-peptides, including differently charged species. These transporters are stereoselective toward peptides containing L-enantiomers of amino acids. Both PEPT1 and PEPT2 exhibit a similar substrate specificity but differ in structure, transport capacity, and binding affinity. PEPT1 is a low-affinity, high-capacity transporter, whereas PEPT2 acts with high affinity and low capacity. Thus, peptide transporters represent excellent targets for the delivery of pharmacologically active compounds because their substrate-binding site can accommodate a wide range of molecules of different size, hydrophobicity and charge.
Metal complexes of peptides have been widely used as models for the interaction between metal ions and proteins, most of these studies concerning Pt(II). Nevertheless, only a few Pt(II)-peptido complexes have been isolated, the major drawback being their kinetic inertness, and only very few studies concerning Au(III)-peptido derivatives with the avowed aim of investigating the direct interaction between Au3+ ion and biomolecules have been reported [M. Wienken, B. Lippert, E. Zangrando, L. Randaccio Inorg. Chem. 1992, 31, 1983-1985; S. Carotti, G. Marcon, M. Marussich, T. Mazzei, L. Messori, E. Mini, P. Orioli, Chem. Biol. Interact. 2000, 125, 29-38].